Correlation between self-reported low back pain and morphological changes in abdominal wall muscles.

BACKGROUND: Morphological changes in the abdominal wall muscle in diastasis recti abdominis (DRA) patients with low back pain (LBP) symptoms and whether there is a wider interrectus distance (IRD) remain unclear. OBJECTIVE: The aim of this study was to determine the correlation between DRA severity and LBP and the morphological differences in abdominal wall muscles between patients who presented with symptoms of LBP and those who were asymptomatic. METHODS: A total of 57 postpartum DRA patients were enrolled, including 21 without LBP, 36 with LBP, and 30 healthy nulliparous women. The numerical rating scale (NRS) was used to evaluate the degree of LBP. The IRD and muscle thickness of the abdominal muscles were measured via an ultrasonography device at the end of a natural breathing cycle. RESULTS: The abdominal muscles of the rectus abdominis (RA), external oblique muscle (EO), and transversus abdominis (TrA) in the DRA patients were significantly thinner than those in the control group (p< 0.05), whereas the difference in the internal oblique muscle (IO) was not significant (p> 0.05). There were no statistically significant differences in abdominal muscle thickness or the IRD regardless of whether the patients with DRA had LBP symptoms, with similar findings between the NRS score and DRA severity (P> 0.05). CONCLUSION: In DRA patients within one year postpartum, no significant difference was found in the severity of DRA or abdominal muscles, regardless of whether they were complicated by LBP symptoms. Shortening the IRD alone may not be beneficial for improving LBP symptoms in DRA patients. The relationship between DRA and LBP and the role of the abdominal muscles in postpartum LBP should be interpreted with caution.

High Degree of Polymerization of Chitin Oligosaccharides Produced from Shrimp Shell Waste by Enrichment Microbiota Using Two-Stage Temperature-Controlled Technique of Inducing Enzyme Production and Metagenomic Analysis of Microbiota Succession.

The direct enzymatic conversion of untreated waste shrimp and crab shells has been a key problem that plagues the large-scale utilization of chitin biological resources. The microorganisms in soil samples were enriched in two stages with powdered chitin (CP) and shrimp shell powder (SSP) as substrates. The enrichment microbiota XHQ10 with SSP degradation ability was obtained. The activities of chitinase and lytic polysaccharide monooxygenase of XHQ10 were 1.46 and 54.62 U/mL. Metagenomic analysis showed that Chitinolyticbacter meiyuanensis, Chitiniphilus shinanonensis, and Chitinimonas koreensis, with excellent chitin degradation performance, were highly enriched in XHQ10. Chitin oligosaccharides (CHOSs) are produced by XHQ10 through enzyme induction and two-stage temperature control technology, which contains CHOSs with a degree of polymerization (DP) more significant than ten and has excellent antioxidant activity. This work is the first study on the direct enzymatic preparation of CHOSs from SSP using enrichment microbiota, which provides a new path for the large-scale utilization of chitin bioresources.

Diastasis Recti Abdominis: A Practical and Effective Width-Length Classification Based on Ultrasound Measurements and its Clinical Validation.

OBJECTIVES: This study aimed to establish a simple and practical classification to guide the clinical treatment of diastasis recti abdominis (DRA) based on ultrasound characteristics with different severities of DRA, and to verify its clinical utility. METHODS: We retrospectively enrolled 301 DRA patients as pilot cohort and divided into Conservative Treatment Group and Surgical Group according to clinical outcomes. A new Width-Length classification was summarized based on ultrasound measurements of the width and length of midline separation. Then, 100 DRA patients were enrolled prospectively as validation cohort, and diagnostic performance was evaluated by clinical treatment. RESULTS: The Width-Length classification in pilot cohort was as follows: Type 1 (n = 108), open only at M3; Type 2 (n = 63), open at M3 and either M2 or M4 (inter-rectus distance at M3 <47 mm); Type 3 (n = 44), open at M3 and either M2 or M4 (inter-rectus distance at M3 ≥47 mm); Type 4 (n = 74), open at M3, along with other two sites of M1, M2, M4, or M5; Type 5 (n = 12), open at M2, M3, and M4, along with M1 or M5, or both. DRA patients in Type 1-2 were recommended for conservative treatment, and in Type 3-5 were recommended for surgical treatment (all P < .05). In the validation cohort, the accuracy of Width-Length classification in determining treatment strategy was 86.0%. CONCLUSIONS: This study proposes a Width-Length classification based on the width and length of midline separation on ultrasound, which was validated to be simple, practical and effective in guiding DRA treatment.

Effects of Laughter Therapy on Improving Negative Emotions Associated with Cancer: A Systematic Review and Meta-Analysis.

INTRODUCTION: With aging and growth of the population, the risk of cancer incidence and mortality is rapidly increasing. However, psychosocial treatment has been seriously neglected in many healthcare settings. Laughter therapy is a therapeutic program to improve emotional wellbeing and health which has been applied as a complementary treatment. We aim to explore effects of laughter therapy for patients with cancer on their negative emotions such as depression, anxiety, stress, pain, and fatigue. METHODS: We searched the Cochrane Library, Embase, PubMed, Scopus, Web of Science, WANFANG data, Weipu (VIP), Chinese National Knowledge Infrastructure (CNKI) and independently rated the risk of bias in every article using the Cochrane Collaboration's Risk of Bias Assessment Tool. Review Manager and STATA software were used to pool the individually included studies. RESULTS: Seven studies were found eligible to be included in the present review. Overall, study quality was relatively high. Our findings suggest that laughter therapy might have a positive effect on improving emotional response in cancer patients. Arguably, laughter therapy, whether humor or laughter, has a positive effect on anxiety, stress, pain feeling, fatigue, and depression in cancer patients. CONCLUSIONS: Laughter therapy is a convenient, multi-modality, flexible-duration therapy to improve negative emotions in cancer patients, regardless of their gender, age, and type of cancer.

Effectiveness of online mindfulness-based interventions for cancer patients: a systematic review and meta-analysis.

PURPOSE: Cancer is the second leading cause of mortality worldwide. Cancer negatively affects individuals' quality of life and overall health. Mindfulness-based interventions appear to be promising in the reduction of cancer- and treatment-related symptoms. This review aimed to determine the effectiveness of online mindfulness-based interventions on distress, anxiety, depression, stress, mindfulness, sleep disturbance, quality of life, rumination, fear of cancer recurrence, fatigue and post-traumatic growth among adult cancer patients. METHODS: A literature search was conducted across five electronic databases. Only randomized controlled trials were eligible. Two reviewers independently screened the studies, extracted data, and performed quality assessment using the Cochrane risk of bias assessment tool. Meta-analyses were conducted using review manager software, and standardized mean difference was used to determine intervention effects. Heterogeneity was examined using the I2 statistics. RESULTS: Ten studies were included with a total of 962 participants. Analyses revealed that online mindfulness-based interventions was effective in reducing distress (I2 = 98%；standardized mean difference = -2.21，95% confidence interval: -3.84 to 0.57；P = 0.008)， depression (I2 = 45%；standardized mean difference = -0.33，95% confidence interval: -0.64 to -0.03；P = 0.03), stress (I2 = 97%；standardized mean difference = -2.14，95% confidence interval: -4.24 to -0.03；P = 0.05) and sleep disturbance (I2 = 54%；standardized mean difference = -0.30，95% confidence interval: -0.59 to -0.01；P = 0.04), and improving quality of life (I2 = 94%；standardized mean difference = 0.92，95% confidence interval: 0.09-1.76；P = 0.03). The online mindfulness-based interventions had no significant effects on anxiety, mindfulness, rumination, fear of cancer recurrence, fatigue and post-traumatic growth. Subgroup analyses revealed that online mindfulness-based interventions resulted in higher effect sizes for distress when delivered by website than application, significantly higher effect sizes were also found for online mindfulness-based interventions with guidance, but not on treatment or cancer type. For sleep disturbance, and quality of life, no significant differences between subgroups were found. CONCLUSION: These results provide preliminary support that online mindfulness-based interventions may be feasible and acceptable, which can be used as an adjuvant therapy for the management of cancer-related symptoms among cancer patients.

A novel lytic polysaccharide monooxygenase from enrichment microbiota and its application for shrimp shell powder biodegradation.

Lytic polysaccharide monooxygenases (LPMO) are expected to change the current status of chitin resource utilization. This study reports that targeted enrichment of the microbiota was performed with chitin by the selective gradient culture technique, and a novel LPMO (M2822) was identified from the enrichment microbiota metagenome. First, soil samples were screened based on soil bacterial species and chitinase biodiversity. Then gradient enrichment culture with different chitin concentrations was carried out. The efficiency of chitin powder degradation was increased by 10.67 times through enrichment, and chitin degradation species Chitiniphilus and Chitinolyticbacter were enriched significantly. A novel LPMO (M2822) was found in the metagenome of the enriched microbiota. Phylogenetic analysis showed that M2822 had a unique phylogenetic position in auxiliary activity (AA) 10 family. The analysis of enzymatic hydrolysate showed that M2822 had chitin activity. When M2822 synergized with commercial chitinase to degrade chitin, the yield of N-acetyl glycosamine was 83.6% higher than chitinase alone. The optimum temperature and pH for M2822 activity were 35°C and 6.0. The synergistic action of M2822 and chitin-degrading enzymes secreted by Chitiniphilus sp. LZ32 could efficiently hydrolyze shrimp shell powder. After 12 h of enzymatic hydrolysis, chitin oligosaccharides (COS) yield reached 4,724 µg/mL. To our knowledge, this work is the first study to mine chitin activity LPMO in the metagenome of enriched microbiota. The obtained M2822 showed application prospects in the efficient production of COS.

New Insights into the Modification of the Non-Core Metabolic Pathway of Steroids in Mycolicibacterium and the Application of Fermentation Biotechnology in C-19 Steroid Production.

Androsta-4-ene-3,17-dione (AD), androsta-1,4-diene-3,17-dione (ADD), and 9α-hydroxy-4-androstene-3,17-dione (9-OHAD), which belong to C-19 steroids, are critical steroid-based drug intermediates. The biotransformation of phytosterols into C-19 steroids by Mycolicibacterium cell factories is the core step in the synthesis of steroid-based drugs. The production performance of engineered mycolicibacterial strains has been effectively enhanced by sterol core metabolic modification. In recent years, research on the non-core metabolic pathway of steroids (NCMS) in mycolicibacterial strains has made significant progress. This review discusses the molecular mechanisms and metabolic modifications of NCMS for accelerating sterol uptake, regulating coenzyme I balance, promoting propionyl-CoA metabolism, reducing reactive oxygen species, and regulating energy metabolism. In addition, the recent applications of biotechnology in steroid intermediate production are summarized and compared, and the future development trend of NCMS research is discussed. This review provides powerful theoretical support for metabolic regulation in the biotransformation of phytosterols.

Therapeutic effect of acupuncture combined montelukast sodium on cough variant asthma in children: A protocol for systematic review and meta-analysis.

BACKGROUND: Cough variant asthma in children is a special type of asthma. Although there are many effective cases of combined acupuncture and western medicine in the clinical treatment of this kind of children, there is no standardized acupuncture combined with western medicine to evaluate the curative effect. Therefore, combined with existing reports, a systematic review and meta-analysis of acupuncture combined with montelukast sodium in the treatment of cough variant asthma in children were carried out to obtain conclusive results. METHODS: The following electronic databases will be searched: PubMed, the Cochrane Library, Embase, Web of Science, Medline, CNKI, Chinese Biomedical Literature Database, VIP, and Wan Fang databases. We will consider articles published between database initiation and October 2021. We will use Review Manager 5.4, provided by the Cochrane Collaborative Network for statistical analysis. Clinical randomized controlled trials related to acupuncture combined with montelukast sodium on cough variant asthma in children were included in this study. Language is limited to both Chinese and English. Research selection, data extraction, and research quality assessments were independently completed by two researchers. We then assessed the quality and risk of the included studies and observed the outcome measures. RESULTS: This study provides a high-quality synthesis to assess the effectiveness and safety of acupuncture combined with montelukast sodium on cough variant asthma in children. CONCLUSION: This systematic review will provide evidence to determine whether acupuncture combined with montelukast sodium is an effective and safe intervention for patients with cough variant asthma in children. INPLASY REGISTRATION NUMBER: INPLASY2021110006.

Cloning, expression and characterization of metalloproteinase HypZn from Aspergillus niger.

A predicted metalloproteinase gene, HypZn, was cloned from Aspergillus niger CGMCC 3.7193 and expressed in Pichia pastoris GS115, and the physicochemical characteristics of recombinant HypZn were investigated after separation and purification. The results showed that the specific activity of the purified HypZn reached 1859.2 U/mg, and the optimum temperature and pH value of HypZn were 35°C and 7.0, respectively. HypZn remained stable both at 40°C and at pH values between 5.0 and 8.0. The preferred substrate of HypZn was soybean protein isolates, and the Km and Vmax values were 21.5 µmol/mL and 4926.6 µmol/(mL∙min), respectively. HypZn was activated by Co2+ and Zn2+ and inhibited by Cu2+ and Fe2+. The degree of soybean protein isolate hydrolysis reached 14.7%, and the hydrolysates were of uniform molecular weight. HypZn could tolerate 5000 mM NaCl and completely lost its activity after 30 min at 50°C. The enzymological characterizations indicated that HypZn has great application potential in the food industry, especially in fermented food processing.

The role of carbonic anhydrase III and autophagy in type 2 diabetes with cardio-cerebrovascular disease.

Type 2 diabetes mellitus (T2DM) is one of the most common chronic diseases among the elderly people. The T2DM increases the risk of cardio-cerebrovascular disease (CCD), and the main pathological change of the CCD is atherosclerosis (AS). Meanwhile, the carbonic anhydrases (CAs) are involved in the formation and progression of plaques in AS. However, the exact physiological mechanism of carbonic anhydrase III (CAIII) has not been clear yet, and there are also no correlation study between CAIII protein and T2DM with CCD. The 8-week old diabetic mice (db/db-/- mice) and wild-type mice (wt mice) were feed by a normal diet till 32 weeks, and detected the carotid artery vascular opening angle using the method of biomechanics; The changes of cerebral cortex and myocardium were watched by the ultrastructure, and the autophagy were observed by electron microscope; The tissue structure, inflammation and cell injury were observed by Hematoxylin and eosin (HE) staining; The apoptosis of cells were observed by TUNEL staining; The protein levels of CAIII, IL-17, p53 were detected by immunohistochemical and Western Blot, and the Beclin-1, LC3, NF-κB were detected by Western Blot. All statistical analysis is performed using PRISM software. Compared with wt mice, db/db-/- mice' carotid artery open angle increased significantly. Electron microscope results indicated that autophagy in db/db-/- mice cerebral cortex and heart tissue decreased and intracellular organelle ultrastructure were damaged. HE staining indicated that, db/db-/- mice' cerebral cortex and heart tissue stained lighter, inflammatory cells infiltration, cell edema were obvious, myocardial fibers were disorder, and myocardial cells showed different degrees of degeneration. Compared with wt mice, TUNEL staining showed that there was obviously increase in db/db-/- mice cortex and heart tissue cell apoptosis. The results of immunohistochemistry and Western Blot indicated that CAIII, Beclin-1 and LC3II/I expression levels conspicuously decreased in cortex and heart tissue of db/db-/- mice, and the expression level of IL-17, NF-κB and p53 obviously increased. The carotid artery' vascular stiffness was increased and which was probably related with formation of AS in diabetic mice. And the autophagy participated in the occurrence and development of diabetic CCD. CAIII protein might somehow be involved in the regulation of autophagy probably through affecting cell apoptosis and inflammation, but the underlying mechanism remains to be further studied.

A new carboxypeptidase from Aspergillus niger with good thermostability, pH stability and broad substrate specificity.

A new serine carboxypeptidase gene, capA, was identified in Aspergillus niger CBS 513.88 by reading genomic information and performing sequence alignment, and the gene was cloned and expressed in Pichia pastoris GS115. In a shake flask, the enzyme activity of the recombinant strain GS115 (pPIC9K-capA) reached 209.3 U mg-1. The optimal temperature and pH for enzyme activity were determined to be 45 °C and 6.0, respectively. After incubation at 40-50 °C or at pH 4.0-8.0 for 1 h, the enzyme retained more than 80% or 60% of its initial activity. The presence of 1-10 mmol L-1 Mg2+ enhanced the activity of CapA, whereas 1-10 mmol L-1 Cu2+, Fe2+, or Co2+, 10 mmol L-1 Mn2+, or 1-10 mmol L-1 phenylmethylsulfonyl fluoride (PMSF) significantly inhibited its activity. CapA had a broad substrate specificity and preferred the hydrophobic amino acids Leu and Lys at the C terminus of proteins, and N-benzyloxycarbonyl-L-phenylalanyl-L-leucine (Cbz-Phe-Leu) was the optimal substrate, for which CapA exhibited Km 0.063 mmol L-1 and kcat/Km 186.35 mmol L-1 s-1. The good thermostability, pH stability and hydrolysis characteristics of CapA provide a solid foundation for application in the food and biotechnology fields.

Coexpression of VHb and MceG genes in Mycobacterium sp. Strain LZ2 enhances androstenone production via immobilized repeated batch fermentation.

Androstenone production is limited by low-efficiency substrate transport and dissolved oxygen levels during fermentation. In this study, the coexpression of the optimized Vitreoscilla hemoglobin (VHb) and sterol transporter ATPase (MceG) genes in Mycobacterium sp. LZ2 (Msp) was investigated to alleviate dissolved oxygen and mass transfer limitations. Results revealed that Msp-vgb/mceG effectively improved the growth, production, and adaptation to dissolved oxygen compared with those of Msp. The increased catalase activity and reduced intracellular ROS levels enhanced cell viability and promoted transcription of genes critical for phytosterol metabolism. Bagasse as an immobilization carrier increased the productivity of Msp-vgb/mceG by 56%. Immobilized repeat batch fermentation reduced the biotransformation period from 60 days to 37 days and improved the productivity from 0.039 g/L/h to 0.069 g/L/h. To the best of our knowledge, this work is the first study on the immobilization of recombinant mycobacteria on bagasse for androstenone production.

Manifestation of acute peritonitis and pneumonedema in scrub typhus without eschar: A case report.

BACKGROUND: Scrub typhus is an acute infectious disease caused by rickettsia infection. The diagnosis is based on eschar, and clinical manifestations can range from asymptomatic to multiorgan dysfunction. CASE SUMMARY: We report the case of a 35-year-old man living in Zhuhai, Guangdong, China, who had repeated high fever with a maximum body temperature of 40.2 °C and elevated white blood cells and procalcitonin levels. After 7 d of persistent high fever, the patient developed rash, abdominal pain, and symptoms of peritonitis. Within 24 h after admission, the patient developed diffuse peritonitis and pneumonedema, requiring ventilator support in the intensive care unit. However, there was no eschar on the body, and the first Weil-Felix test was negative. Taking into account that the patient had a history of jungle activities, doxycycline combined with meropenem was selected. The patient improved, healed, and was discharged after a week. The diagnosis of scrub typhus was confirmed by a repeat Weil-Felix test (Oxk 1:640), and pathology of the appendix resected by laparotomy suggests vasculitis. CONCLUSION: This rare presentation of peritonitis, pulmonary edema, and pancreatitis caused by scrub typhus reminds physicians to be alert to the possibility of scrub typhus.

Serum Heparanase: A New Clinical Biomarker Involved in Senile Metabolic Inflammatory Syndrome.

AIM: Metabolic inflammation syndrome (MIS) can lead to a series of complications, but its exact inflammatory mechanism is still unclear. The aim of this study was to explore the correlation between heparanase (HPA) and MIS, and the close relationship between HPA and other chronic low-grade inflammation index, such as C-reactive protein (CRP) and interleukin-6 (IL-6). METHODS: A total of 105 patients with MIS in the physical examination population of Huashan Hospital affiliated to Fudan University from May to June 2018 were selected as the MIS group, and 52 patients who were relatively healthy during the same period were used as the control group. The basic clinical data of the selected candidates were collected, the levels of serum HPA, CRP and IL-6 were measured by ELISA, and the levels of blood glucose and blood lipids were also detected. RESULTS: Compared with the control group, the levels of HPA, CRP, IL-6, FBG, HbA1C, and TG of MIS group were all significantly elevated (all P<0.05), and HDL-C levels were considerably reduced (P<0.05). Correlation analysis showed that there was a noticeably positive correlation between serum HPA level and CRP, IL-6 levels (P<0.05). CONCLUSION: Higher HPA levels might play a certain role in the occurrence and development of MIS. There was a certain close correlation between serum HPA level and CRP and IL-6 levels, and which indicated that HPA was involved in the chronic low-grade inflammatory reaction process of MIS.

Muscle elasticity is different in individuals with diastasis recti abdominis than healthy volunteers.

OBJECTIVE: To determine the value of shear wave elastography (SWE) in assessing abdominal wall muscles, including rectus abdominis (RA), external oblique muscle (EO), internal oblique muscle, and transversus abdominis (TrA) in patients with diastasis recti abdominis (DRA) and healthy controls. METHODS: From October 2018 to December 2019, 36 postpartum DRA patients and 24 nulliparous healthy women were identified. Inter-rectus distance (IRD) measurements were taken by B-mode ultrasound. Shear wave speed (SWS) values were acquired by one operator at ten specific locations. Clinical and ultrasound variables, including demographics, IRD, muscle thickness, and muscle SWS, were compared between the two groups using Student's t test or Fisher's exact test. Pearson correlation analyses were conducted for the variables of IRD, muscle thickness, and SWS in the 36 DRA patients. RESULTS: The maximum diameter of recti abdominus separation was located at the umbilicus in DRA patients (4.59 ± 1.14 cm). The SWS value was significantly lower in the RA (p = 0.003) and higher in the TrA muscle (p < 0.001) in DRA patients compared with the age-matched controls. However, SWS in both muscles (RA and TrA) showed a statistically positive correlation with IRD (p < 0.05). In addition, the SWS value in EO statistically decreased in DRA patients compared with the healthy controls (1.65 ± 0.15 vs. 1.79 ± 0.14, p = 0.001). CONCLUSIONS: The application of SWE to abdominal wall muscles in DRA patients is feasible. The correlation between SWS value and IRD in RA should be interpreted with caution.

Combined carbon photon and hydrogel therapy mediates the synergistic repair of full-thickness skin wounds.

OBJECTIVE: This study investigated the synergistic repair effects of Prontosan hydrogel and carbon photon therapy in a rat full-thickness wound model. METHODS: The wavelength distribution of the photon source was determined. Dehydration of the Prontosan hydrogel and fibroblast viability were analyzed following exposure to different durations of light exposure at different distances from the source. Indexes of wound healing in a full-thickness rat wound model were then determined in groups (n = 8 each) subjected to either no treatment, Prontosan treatment only, carbon photon therapy only, or a combination of the two treatments (synergistic group). RESULTS: Carbon photon exposure for 15 minutes at a distance of 20 cm from the wound was found to be optimal. Wound healing occurred faster in the synergistic group compared with the control and single-treatment groups. Growth factor secretion, granulation tissue formation, inflammation regulation, collagen deposition, and neovascularization were all higher in the synergistic group. CONCLUSIONS: Prontosan hydrogel combined with carbon photon therapy may provide an optimal environment for wound healing and serve as a novel physical approach to the treatment of wounds. However, the number of animals included in this study was relatively small and a larger study is required to confirm these findings.

Efficient repeated batch production of androstenedione using untreated cane molasses by Mycobacterium neoaurum driven by ATP futile cycle.

The biotransformation of phytosterol to androstenedione (AD) by mycobacteria is a unique process accompanied by energy-producing. However, high intracellular ATP content can severely inhibit the efficient production of AD. In this study, a novel citrate-based ATP futile cycle (AFC) and pyruvate-based AFC were constructed for the first time. Application of AFCs reduced intracellular ATP and propionyl-CoA levels and increased NAD+/NADH ratios and cell viability. The forced consumption of ATP promotes the transcription of critical genes in propionyl-CoA metabolism. The synergistic effect of enhanced propionyl-CoA metabolism and AFC increased AD conversion yield from 60.6% to 97.3%. The AD productivity was further improved by repeated batch fermentation using untreated cane molasses. The maximum productivity was 181% higher than that of the original strain. Therefore, the strategy of combining AFC and repeated batch fermentation is a valuable tool for the efficient and low-cost production of AD and other steroidal pharmaceutical precursors.

Improving phytosterol biotransformation at low nitrogen levels by enhancing the methylcitrate cycle with transcriptional regulators PrpR and GlnR of Mycobacterium neoaurum.

BACKGROUND: Androstenedione (AD) is an important steroid medicine intermediate that is obtained via the degradation of phytosterols by mycobacteria. The production process of AD is mainly the degradation of the phytosterol aliphatic side chain, which is accompanied by the production of propionyl CoA. Excessive accumulation of intracellular propionyl-CoA produces a toxic effect in mycobacteria, which restricts the improvement of production efficiency. The 2-methylcitrate cycle pathway (MCC) plays a significant role in the detoxification of propionyl-CoA in bacterial. The effect of the MCC on phytosterol biotransformation in mycobacteria has not been elucidated in detail. Meanwhile, reducing fermentation cost has always been an important issue to be solved in the optimizing of the bioprocess. RESULTS: There is a complete MCC in Mycobacterium neoaurum (MNR), prpC, prpD and prpB in the prp operon encode methylcitrate synthase, methylcitrate dehydratase and methylisocitrate lyase involved in MCC, and PrpR is a specific transcriptional activator of prp operon. After the overexpression of prpDCB and prpR in MNR, the significantly improved transcription levels of prpC, prpD and prpB were observed. The highest conversion ratios of AD obtained by MNR-prpDBC and MNR-prpR increased from 72.3 ± 2.5% to 82.2 ± 2.2% and 90.6 ± 2.6%, respectively. Through enhanced the PrpR of MNR, the in intracellular propionyl-CoA levels decreased by 43 ± 3%, and the cell viability improved by 22 ± 1% compared to MNR at 96 h. The nitrogen transcription regulator GlnR repressed prp operon transcription in a nitrogen-limited medium. The glnR deletion enhanced the transcription level of prpDBC and the biotransformation ability of MNR. MNR-prpR/ΔglnR was constructed by the overexpression of prpR in the glnR-deleted strain showed adaptability to low nitrogen. The highest AD conversion ratio by MNR-prpR/ΔglnR was 92.8 ± 2.7% at low nitrogen level, which was 1.4 times higher than that of MNR. CONCLUSION: Improvement in phytosterol biotransformation after the enhancement of propionyl-CoA metabolism through the combined modifications of the prp operon and glnR of mycobacteria was investigated for the first time. The overexpress of prpR in MNR can increase the transcription of essential genes (prpC, prpD and prpB) of MCC, reduce the intracellular propionyl-CoA level and improve bacterial viability. The knockout of glnR can enhance the adaptability of MNR to the nitrogen source. In the MNRΔglnR strain, overexpress of prpR can achieve efficient production of AD at low nitrogen levels, thus reducing the production cost. This strategy provides a reference for the economic and effective production of other valuable steroid metabolites from phytosterol in the pharmaceutical industry.

Economical production of androstenedione and 9α-hydroxyandrostenedione using untreated cane molasses by recombinant mycobacteria.

Production of androstenedione (AD) and 9α-hydroxyandrostenedione (9α-OH-AD) by recombinant mycobacteria using untreated cane molasses and hydrolysate of mycobacterial cells (HMC) was investigated for the first time. B-vitamins feeding experiment and reverse transcription-PCR analysis showed that propionyl-CoA carboxylase (PCC) plays an important role in the phytosterol biotransformation of mycobacteria. The respective AD and 9α-OH-AD conversion ratios were increased by 2.91 and 1.48 times through coexpression of PCC and NADH dehydrogenase. The highest conversion ratios of AD and 9α-OH-AD obtained by using a co-feeding strategy of cane molasses and HMC reached 96.38% and 95.04%, respectively, and the total costs of carbon and nitrogen sources for the culture medium were reduced by 29.89% and 49.49%, respectively. Taking the results together, untreated cane molasses and HMC can be used for the economical production of steroidal pharmaceutical precursors by mycobacteria. This study offers an economical and green strategy for steroidal pharmaceutical precursor production.